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    • lysophosphatidic acid The electrical properties of cells wer

    2018-10-26

    The electrical properties of lysophosphatidic acid were recently analyzed by EIS during the differentiation of stem cells (Angstmann et al., 2011; Bagnaninchi and Drummond, 2011; Venkatanarayanan et al., 2013). These pioneering works have demonstrated a possibility of using changes in impedance as a biomarker for stem cell behaviors. However, the studies were limited to qualitative analyses, unable to fully realize the quantitative potentials of EIS to correlate the physical (cell quantity and morphology) and electrical (resistive and capacitive) properties of the cells with stem cell self-renewal/differentiation.
    Results
    Discussion During stem cell differentiation, the cells undergo transient changes in cell shape until reaching end-phenotypes. These physico-morphological changes are important markers to determine the differentiation state of the cells (Neuhuber et al., 2004; Sullivan et al., 2010). Optical microscopy has been a choice of non-destructive analytical methods to monitor cellular behaviors. However, an optical analysis is semi-quantitative at best, and often subjective by observers. In this regard, a new quantitative analytical methodology enabling real-time observation of cellular behaviors in a non-destructive manner would advance our fundamental understanding in stem cell biology. Recently, Raman spectroscopy demonstrated its ability to assess the degree of stem cell differentiation by non-destructively detecting macromolecular compositional changes (Chan et al., 2009; Konorov et al., 2015; Schulze et al., 2010). However, such chemical changes are associated with protein expression, which typically occur at the later stages of stem cell differentiation. In this study, we demonstrated that the differentiation lineage/state-dependent morphological changes of human IPSCs were significantly correlated to the changes in mass and impedance spectra that were continuously monitored throughout the culture period, providing a quantitative means to determine stem cell development. Monitoring the mass changes provides information about general cell expansion, but it alone cannot completely depict cellular behaviors especially after reaching 100% confluency. For example, there was an increase in mass corresponding to an increase in surface coverage up 100% coverage at hour 60. Following hour 60, there were differences in mass change patterns, yet the surface coverage remained at 100%, due to changes in cell size. In this regard, impedance measurement can supplement the missing information, i.e., cell morphology, as impedance response depends on the electrical current pathway determined by cell surface coverage and cell morphology (Giaever and Keese, 1986, 1993; Keese and Giaever, 1994; Wegener et al., 2000). As expected, impedance measurements were similar at hour 48 before the cells reached 100% surface coverage at hour 60, regardless of their culture conditions. Thereafter, differential and dynamic impedance changes were observed among the conditions, likely due to the combinational effects from lineage-/stage-dependent cell morphology changes. Interestingly, after the cells undergoing ectodermal differentiation reached 100% confluency, up to which similar mass and impedance changes to the self-renewal condition were observed, there was a differential development in impedance spectrum. Typically, cells subjected to ectodermal differentiation become tightly packed to form neural rosettes (Elkabetz et al., 2008; Warren et al., 2010). These tightly packed cells stack on top of each other, therefore affecting the electrical pathway, leading to possible increase in impedance at lower frequencies. In contrast, cells undergoing mesendodermal differentiation exhibited a slightly different development of impedance spectrum from the initiation of differentiation compared with the other conditions. This is likely due to the changes in cell shape before reaching 100% confluency. The changes in cell size and morphology continued with a mass decrease while maintaining 100% surface coverage. This resulted in fewer cell-cell junctions, which may have led to the changes in impedance. However, it is not readily possible to decouple the individual contributions of such changes in cell coverage, shape, and cell-cell/cell-substrate interactions from the overall impedance of the complex system.